1. Field of the Invention
The present invention relates to a method of assay of a biological associated material. The present invention also relates to a reaction vessel used in the assay of the biological associated material.
2. Description of the Related Art
Biological associated materials (i.e., material contained in a body fluid) have been assayed as daily examinations in the medical and environmental sanitation fields. Particularly, in the medical field, various types of biological associated materials have been assayed in many medical centers to diagnose diseases and to evaluate therapeutic effects for diseases. Examples of the biological associated material are biological constituent materials such as various proteins (including antibodies), polypeptides, saccharides, nucleic acid, lipids, and various hormones represented by steroid, in addition to extravital materials such as various pathogenic microorganisms, an allergen for causing an allergic reaction, and drugs for causing biological reactions. The particular materials having a relevance to generation of contagious diseases or cancer and cause an antigen-antibody reaction or a reaction between a nucleic acid and another complementary nucleic acid.
Such a biological associated material is assayed in a reaction with a specific affinity material having a specific affinity with this biological associated material. For example, using an antigen-antibody reaction as one of the reactions between a biological associated material and a corresponding specific affinity material, infectious diseases such as acquired immunodeficiency syndrome (to be referred to as AIDS hereinafter) as the most critical current topic in public health can be examined, and cancer associated materials which can hardly be conventionally specified can be assayed.
A DNA or RNA as a gene of an infectious microorganism can also be assayed using a complementary nucleotide which binds to the characteristic portion of the polynucleotide of the DNA or RNA. The reaction between the polynucleotide and complementary nucleotide is one of the reactions between the biological associated materials and the corresponding specific affinity materials. A reaction between insulin as one of hormones and an insulin receptor is one of the reactions between the biological associated materials and the corresponding specific affinity materials.
In this manner, various methods of assaying biological associated materials using reactions with specific affinity materials are known. In any method, the amount of a biological associated material (to be referred to as a bound material hereinafter) to which a specific affinity material is bound must be assayed.
These methods of assaying the biological associated materials are mainly classified into two methods. The first method is an assay for detecting a bound material by utilizing a change in nature of a specific affinity material itself or a tracer combined thereto upon binding of the specific affinity material to the bound material. This first method has been called a homogenous assay. The second method is an assay for performing B (Bound)/F (Free) separation for separating a bound material from a free material after a complex of a specific affinity material and a bound material is set insoluble by any means. The second method has been called a heterogeneous assay.
As the heterogeneous assay, a method is also known in which a second specific affinity material which specifically binds to a complex of a specific affinity material and a bound material is bound to the complex so that the size of the complex molecule is increased, thereby setting the complex insoluble. This method, however, is not preferable due to poor reliability in assay precision. In order to solve this problem, the following B/F separation is generally performed. A specific affinity material is bound to an insoluble material. Thereafter, the specific affinity material is reacted with a biological associated material so as to form a complex of the biological associated material, the specific affinity material and the insoluble material, and then the complex is separated from a sample solution. Examples of the insoluble material are a reaction tube, beads, or filter paper.
In the heterogeneous assay, when a reaction tube itself is used as an insoluble material, the reaction tube is used only once and need not be cleaned, thus requiring a relatively small amount of cleaning water. However, waste products tend to increase in amount, resulting in inconvenience. This assay is not suitable for treating a large number of samples. In addition, the substantial reaction between the biological associated material and the specific affinity material is assumed to occur on the surface of the reaction tube. For this reason, a sample and a reagent which cannot be reacted with the specific affinity material are presented, and are wasted, resulting in inconvenience.
To the contrary, when a material such as beads or filter paper except for the reaction tube is used as an insoluble material, the reaction tube can be repeatedly used. However, since the reaction tube must be cleaned, an assay operation becomes cumbersome. If the operation is automated, the resultant apparatus becomes bulky. The sample to be examined tends to be left (so-called carry-over) in the insoluble material.
In addition, the precision and efficiency of heterogeneous assay depend on whether B/F separation is effectively performed. When the reaction tube is used as an insoluble material, the reaction tube can be relatively easily separated from a sample solution. To the contrary, when an insoluble material except for the reaction tube is used, for example, a filter is used to separate the insoluble material from the sample solution, or an insoluble material is agglutinated and separated by a centrifugal or magnetic force. For this reason, it is difficult to precisely perform B/F separation at high speed.
As a means for solving the disadvantage of the conventional heterogeneous assay described above, the present applicant proposes a novel method for assay of a biological associated material (U.S. Pat. No. 5,066,465). A reaction vessel used in this method has a sample inlet channel having a sectional area capable of drawing a sample in the reaction vessel by a capillary phenomenon, a recess formed in the inner wall of the sample inlet channel, and a transparent plate disposed above the recess and having a flat surface for defining the upper limit of a reaction region. In this method, a sample is dripped in the sample inlet channel to cause a predetermined amount of the sample to be absorbed in the reaction region by the capillary phenomenon, and a pattern of agglutinated particles is formed in the recess, thereby analyzing the biological associated material. This method is suitable for an assay using a very small amount of a sample.
Since a reaction between a biological associated material and a specific affinity material generally requires a reaction time of minutes to tens of minutes, a sample solution must be caused to stay in the reaction vessel for a predetermined period of time. On the other hand, in cleaning the reaction vessel, the cleaning solution unlike the sample solution need not stay in the reaction vessel and must be quickly removed. As a method of assay of a biological associated material using the above reaction vessel, it has been desired earnestly to develop a new method of efficiently holding a solution in a reaction vessel and removing the solution from the reaction vessel so as to perform a more efficient operation.
In order to satisfy the above demand, an evacuation unit is arranged at one end of the sample inlet channel to draw the cleaning solution in the reaction vessel having the above arrangement. However, to perfectly clean the reaction vessel, the cleaning solution must be continuously supplied to the reaction vessel for a predetermined period of time. For this purpose, a cleaning solution supply unit capable of continuously supplying the cleaning solution must be arranged at the other end of the sample inlet channel. Although the reaction vessel itself can be made compact, the resultant apparatus becomes bulky. As a result, a method of assay of a biological associated material using a compact apparatus in a small test center cannot be provided.